(117f) Comparison of Large-Pore Media and Monoliths for Protein and VLP Chromatography

Authors: 
Carta, G., University of Virginia

In this paper we examine the performance of large-pore particles with that of monoliths for the cation exchange chromatography of proteins and VLPs. Chromatographic experiments in bind-and-elute mode are conducted with POROS HS 50 and HS 20, having mean particle size in the 50 and 20 um range, respectively, from Applied Biosystems, and with radial flow monoliths, 1 mL in volume, from BIA Separations. Structural characteristics of both particles and monoliths are determined by scanning and transmission electron microscopy as well as by inverse size exclusion chromatography. The particles are found to have much higher equilbrium binding capacities than the monoliths for proteins, including IgG and thyroglobulin. The dynamic binding capacities are also higher for particle based columns except at extremely high flow rates exceeding 1000 cm/h. Protein recoveries are high for both particles and monoliths. In the case of VLPs, binding is limited to a thin layer on the outside surface of the particles, so that the capacity advantage of the POROS resins is greatly reduced compared to the monoliths. However, VLP recovery from the monoliths is low and is dependent on the direction of flow. Closer microscopic examination, indicates that physical heterogenities in the monolith structure may result in physical trapping of the VLPs.